There is not a single FDA-approved drug that halts or even slows neurodegeneration in the CNS. Despite spending billions of dollars, neither industry nor academia has been able to develop a single drug that slows the progression of Alzheimer's (AD), Parkinson's (PD) and Creutzfeldt-Jakob (CJD) diseases as well as amyotrophic lateral sclerosis (ALS) and fronto-temporal dementia (FTD). Despite this roadblock, there have been impressive advances in understanding the pathogenesis of all these disorders. A steady accumulation of experimental data argues that a different protein causes each neurodegenerative disease and these proteins acquire alternative structures that become self-propagating i.e., prions (Meyer-Luehmann et al., 2006; Clavaguera et al., 2009; Frost and Diamond, 2009; Olanow and Prusiner, 2009; Brundin et al., 2010; Cushman et al., 2010; Colby and Prusiner, In press). These findings are most gratifying since they provide an enlarging body of evidence in support of what now are regarded as prescient speculations (Prusiner, 1984, 2001). The understanding that all or most ofthe neurodegenerative diseases are caused by prions may give several new perspectives on the development of effective therapeutics. First, drugs that cure cultured cells infected with prions may not predict success in experimental animals or humans. Such is our experience with the antimalarial drug quinacrine that cured cultured cells but failed to extend the lives of either mice or humans (Collinge et al., 2009; Ghaemmaghami et al., 2009). Even when the level of quinacrine was increased almost 100-fold, in the brains of mice in which the P-gp transporter (Mdr1) has been knocked out, above the halfeffective concentration (EC50) in cultured cells, it failed to extend the incubation time (Ghaemmaghami et al., ^ 2009). We were able to gather convincing evidence showing that the most likely explanation for this therapeutic failure was a conformational change in PrP^*^ resulting in a drug-resistant prion strain. These results prompted us to develop a broad drug discovery program that has begun to identify lead compounds that are able to extend incubation times in mice (Ghaemmaghami et al., 2010; Gallardo-Godoy et al., 2011). Discovering drugs that can be used to treat neurodegeneration presents substantial challenges. With that said, we desperately need additional assays for measuring the efficacy of hits and leads to proceed through the process of drug discovery. More predictive assays of lead efficacy in cultured cells prior to studies in Tg rodents are critical to the development of effective therapeutics. Such cell assays could save an immense amount of time and resources. To improve our in vitro assessments of chemical libraries as well as groups of lead compounds, we plan to use a newly acquired Opera confocal microscope system for high throughput screening (HTS). This advanced system will allow us to measure both PrP^ and PrP^'^ in subcellular compartments. Despite successfully adapting ELISAs for measuring these PrP isoforms in HTS, we anticipate that the robust resolution of the Opera system will allow us to predict more accurately, from cell-based assays, which compounds will advance successfully through animal models. The adaption of bioluminescence to the in vivo readout of experimental scrapie and Alzheimer's disease in Tg mice (Tamgtiney et al., 2009a; Watts et al., 2011) has greatly facilitated our drug efficacy studies. Plans for improving these mouse models and extending the work into rats are described below in the research plan. Bigenic mice-on the Mdrl knockout background and with a luciferase reporter expressed under control of the Gfap promoter-are being bred for use in drug efficacy studies. Similar mice expressing luciferase are being bred, in which the mouse PrP gene has been knocked out and a chimeric human (Hu)/mouse (Mo) transgene is also expressed. Such mice become ill <80 days after inoculation with CJD(MM1) prions (Giles et al., 2010). In another set of studies, we have used RNAi libraries to identify auxiliary proteins that participate in the formation and replication of PrP?*^ prions. These studies provide a route into the identification of new drug targets for antiprion drugs. More emphasis on these studies is planned since it seems likely that cocktails of drugs with different modes of action will be the most likely routes to successful treatment of neurodegenerative illnesses.